The transformation from caterpillar to butterfly or moth is one of the most beguiling in the animal world. Both larva and adult are just stages in the life of a single animal, but are nonetheless completely separated in appearance, habitat and behaviour. The imagery associated with such change is inescapably beautiful, and as entrancing to a poet as it is to a biologist.

According to popular belief, within the pupa, the caterpillar’s body is completely overhauled, broken down into a form of soup and rebuilt into a winged adult. Richard Buckmister Fuller once said that “there is nothing in a caterpillar that tells you it’s going to be a butterfly.” Indeed, as the butterfly or moth quite literally flies off into a new world, it is tempting to think that there is no connection between its new life and its old existence as an eating machine.

But not so. A new study has provided strong evidence that the larval and adult stages are not as disparate as they might seem. Adult tobacco hookworms – a species of moth – can remember things that it learned as a caterpillar, which means that despite the dramatic nature of metamorphosis, some elements of the young insect’s nervous system remain intact through the process.

Training caterpillars

Using some mild electric shocks, Douglas Blackiston from Georgetown University trained hookworm caterpillars (Manduca sexta) to avoid the scent of a simple organic chemical – ethyl acetate. The larvae were then placed in the bottom end of a Y-shaped tube, with the scent of ethyl acetate wafting down one arm and fresh air coming down the other. Sure enough, 78% of the trained caterpillars inched down the odour-free arm.

As the caterpillar moulted their way through the larval stage, their aversion to ethyl acetate remained. Blackiston allowed them to pupate and emerge as full-grown moths, before testing them again, about a month after their initial ‘electric’ education. Bear in mind that a tobacco hornworm lives for about 30 to 50 days, so a month is very close to its entire lifespan.

Amazingly, 77% of the adult moths also avoided the ethyl acetate-scented arm of the Y-shaped tube and the vast majority of these were the adult versions of the same larvae that had correctly learned the behaviour originally. Clearly, the larvae had learned to avoid the chemical and that memory carried over into adulthood.

Even so, Blackiston was careful to rule out alternative explanations. For a start, ethyl acetate isn’t naturally foul-smelling. It’s actually rather reminiscent of pear drops and when larvae are exposed to it in the absence of electric shocks, neither they nor the adults they become learn to avoid it.

Another possible explanation hinges on the fact that adults emerging from the pupa usually experience a similar milieu of smells to their caterpillar selves. This chemical legacy’ could explain why adults and larvae react similarly to some odours. But when Blackiston applied ethyl acetate gel to the pupae of untrained caterpillars, the adults did not shrink away from the chemical. Nor did washing the pupae of trained caterpillars, to get rid of any lingering traces of ethyl acetate, have any effect.

How and why?

Blackiston was convinced that some aspect of the caterpillar’ nervous system was carried over into adulthood. However, he also found that this only happened if caterpillars are trained at the last possible stage before they pupate – the ‘fifth instar’. Any earlier, and the memories don’t stick.

The fruitfly Drosophila suggests why this might happen. In its brain, memories of smells are located in mushroom bodies, brain structures that consist of three lobes. The gamma lobe develops very early while the alpha and beta lobes develop just before the pupal stage.

Blackiston thinks that long-lasting larval memories are writ into the alpha and beta lobes, whose neural networks are kept around while the rest of the caterpillar breaks down. If the larvae are too young, these areas haven’t developed yet and any learned information is stored in the gamma lobe and lost when its connections are trimmed back in the pupa.

But why bother? After all, the entire advantage of metamorphosis rests on the very different lifestyles and habitats of caterpillars and moths, which allow them to avoid competing with each other. Nonetheless, moths and butterflies must still return to the right sort of plant in order to lay their eggs and Blackiston suggests that their larva-hood memories may help them to do so.

Comments

This reminds me of the astonishing experiment in which one trains flatworms to change their normal light avoidance behaviour with a punishment/reward. These worms are ground up and fed to their untrained commrades…who then display the same behaviour without having to learn from scratch. Somehow the chemical signals can be ‘eaten’ and absorbed. This made quite an impression on me as a young biology student and contributed to my decision to pursue molecular biology research. I hope that young people reading this review by Ed Young on of the article on Caterpillar metamorphosis are similarly inspired.

You got to the response before I could Ed. Geoffrey, I can back up what he has already said by saying most of the “cannibalism” learning experiments are thought to have been confounded in either execution or interpretation.

However, I have heard some exciting preliminary data from a flatworm lab looking at metamorphosis. Flatworms can regenerate really well, if you cut them in half the front portion grows a new tail and the back portion grows a new head (and in nature many species actually pull themselves in half to reproduce).

What the lab has been looking at is whether a trained flatworm can remember information after regenerating. So far they have shown that if the head regenerates a tail, the worm remembers, not surprising since the “brain” is in the front end. But, what might be very exciting is that when the tail half regenerates a head, it may also remember the learned behavior. The work is very preliminary, but I thought it might be interesting to think about.

There’s a lot of experiments you could do on that front. Instead of cutting the worm in half at the center, or waist, why not cut it into right and left halves. If each side has half a brain, would they remember a learned task after regenerating? What could this tell us about the laterality of information storage?

The point is well made! Observer bias is the nemesis of scientific progress!
The flatworm experiment is an example forcing observation to support an hypothesis. In the case the hypothesis…that complex behavioral patterns can be shared or triggered by simple molecules…has been born out by the body of research on nematodes. The neurological and biochemical circuits of these simple animals can be easily reprogrammed by feeding them regulatory small RNA. The ability to regulate gene expression in nematodes by this mechanism has allowed researchers to rapidly turn on and off gene expression contributing to our understanding of gene regulation in eukaryotes.
So a hypothesis is not proved until it is repeatedly tested.. Nor is it disproved so easily. Let this be a lesson to those that would believe otherwise.

Yes! We did this very experiment in high school biology class for our honors project. We trained avoidance of darkness by shocking worms when they crossed a barrier to dark. This avoidance behavior is opposite to the normal programming of the worm. After carefully resecting the animals one side derived from the original animal of the maintained the training 70% of the time. There was no statistically significant avoidance for regenerated untrained animals. It would be interesting to follow this up with modern technical approaches. Are these inherited wiring paths, chemical gradients that persist, or small molecule behavior triggers?

One other note.. at about the same time the flatworm experiments were taking off, genetics began to show how complex organisms could be encoded in a single strand of DNA. In a bit of a stretch, it was suggested that the sequence of RNA could encode memory. Now we understand that the details of this hypothesis were naive. Rather, RNA can serve as a transmissible trigger that can turn off or on genetic and metabolic circuits triggering relatively complex outcomes. So it isn’t possible for a human cannibal to learn a classmates term paper or for a mouse to learn a maze by eating the brain of another that has. But it is conceivable to transmit small signaling molecules between simpler animals that can activate or inhibit latent inherent pathways in another.